During the previous funding period, we made a number of fundamental observations relative to the mechanism of action of estrogens on mitochondria that guide the aims of the present proposal. We have evidence that the bioenergetic crisis seen during normal brain aging and in AD is caused by mitochondrial structure, function and mobility dysfunctions that leads to a breakdown in synaptic integrity resulting in cognitive decline that characterizes both aging and AD. The present continuation of this grant will further assess the mechanism(s) of effects of estrogens on mitochondria and determine if these effects occur in vivo and in post-mortem samples from women. We will address 4 specific aims. Specific Aim 1 will determine if pharmacological antagonism or genetic reduction in the PKA/DRP1 pathway leads to a loss of synaptic integrity, mitochondrial fission and immobility, and bioenergetic decline in primary hippocamal neurons. Specific Aim 2 will determine if ovariectomy for 2, 12 or 20 weeks compromises the PKA/DRP1 pathway leading to synaptic loss and mitochondrial dysfunction and if these deficits can be restored by E2, an ER(3 agonist, DPN, or P4 treatment for 6 weeks, in vivo. Specific Aim 3 will determine if age and post-ovariectomy duration, changes the synaptoneurosome response to E2, DPN or P4. Specific Aim 4 will determine if therapy with DPN improves PKA/DRP1 pathway function, thereby ameliorating loss of synaptic integrity, mitochondrial immobility and fragmentation seen in a 5XFAD mice model. For all of the aims, we will assess DRPI phosphorylation state, a panel of pre- and post-synaptic markers, and a panel of bioenergetic measures. For aims 1 and 4, we will conduct a detailed assessment of mitochnodrial fragmentation and mobility. Successful completion of these proposed studies could lead to new understanding of estrogen targets in the brain as well as potential new therapies for age-related cognitive decline and AD.